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Title: A 454 sequencing approach to dipteran mitochondrial genome research

Author
item MEGANATHAN, P - Mississippi State University
item SINGH, BANESHWAR - West Virginia University
item WELLS, JEFFREY - West Virginia University
item Guerrero, Felicito
item RAY, DAVID - Mississippi State University

Submitted to: Genomics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/5/2014
Publication Date: 11/11/2014
Publication URL: http://handle.nal.usda.gov/10113/61888
Citation: Meganathan, P.R., Singh, B., Wells, J.D., Guerrero, F., Ray, D.A. 2014. A 454 sequencing approach to dipteran mitochondrial genome research. Genomics. 105:53-60.

Interpretive Summary: Diptera, one of the largest metazoan orders, comprises ~ 150,000 known and/or undescribed species and accounts for 10- 15 % of animal diversity. The group harbors many forensically, medically and agriculturally important flies. However, genetic data available for Diptera is limited when compared to the diversity of the group. As a result, many aspects of dipteran evolution, genetics and phylogenetic relationships remain unclear. Mitochondrial genomes and their genes are very useful to study evolutionary history and relationships between species. These genomes are also utilized in forensic entomology, a discipline of forensic science where the information about insects is used in crime scene investigation, often to estimate post-mortem intervals. Unfortunately, the availability of complete mitochondrial genomes for this group is sparse. Of ~150,000 known species, only 51 complete mitochondrial genomes are available for Diptera in the public databases. In this study, we used next generation high throughput DNA sequencing methods to generate mitochondrial sequence data for four flies, Cochliomyia hominivorax, Haematobia irritans, Phormia regina and Sarcophaga crassipalpis. Our results show that next generation methodology is an efficient method of obtaining complete mitochondrial genomes for dipteran species.

Technical Abstract: The availability of complete mitochondrial genome data for Diptera, one of the largest Metazoan orders, in public databases is limited. Herein, we generated the complete or nearly complete mitochondrial genomes for Cochliomyia hominivorax, Haematobia irritans, Phormia regina and Sarcophaga crassipalpis using a second generation sequencing approach, 454 sequencing technology. We assembled 15,094 bp, 15,043 bp, 15,513 bp, and 15,420 bp mitochondrial genome sequences for C. hominivorax, H. irritans, P. regina and S. crassipalpis, respectively. Comparisons between newly obtained and existing assemblies for C. hominivorax and H. irritans revealed no major discrepancies between versions and verified the utility of 454 sequencing for dipteran mitochondrial genomes. We also report, for the first time, the complete mt-genome sequences for two forensically important fly species, P. regina and S. crassipalpis which could be used in the field of forensic entomology for providing useful information to legal personnel. Comparative analyses of dipteran mitochondrial genomes revealed that all the dipteran species follow similar codon usage and nucleotide biases which could be due to strong mutational and weak selection pressures. This study illustrates the utility of a 454 sequencing approach to obtain complete mitochondrial genomes for dipterans without aid of conventional molecular techniques and validates this method of mt-genome sequencing in arthropods.